PIRIMIPHOS-METHYL
First draft prepared by D.J. Clegg
Carp, Ontario, Canada
EXPLANATION
Pirimiphos-methyl was previously evaluated by JMPR in 1974 and
1976 (Annex 1, references 22 and 26). An ADI of 0-0.01 mg/kg bw was
established in 1976. Relevant data from the previous monograph and
monograph addendum are incorporated into the present monograph.
EVALUATION FOR ACCEPTABLE DAILY INTAKE
Biochemical Aspects
Absorption, distribution, and excretion
Rats
Oral administration of 0.6 mg 2-14C-ring labelled pirimiphos-
methyl/kg bw to 5 male rats resulted in a mean urinary excretion of
80.7% and mean faecal excretion of 7.3% in 24 h, indicating rapid
absorption. At 96 h, 86.0% and 15.2% of the administered dose had
been excreted in urine and faeces, respectively. Nine metabolites
(unidentified) were present in the urine (Bratt & Dudley, 1970).
Female rats given 7.5 mg 2-14C pirimiphos-methyl/kg bw orally
were bled (cardiac puncture, 3 rats per time interval) at 0.5, 1, 3,
5, 7 or 24 h post-dosing. Maximum blood levels (at 0.5 h) were 2-3
µg/ml, declining by 50% 1 h post-dosing. By 24 h, levels of 14C in
blood were 0.2 to 0.3 µg/ml, and of pirimiphos-methyl, 0.01-0.02
µg/ml. Rats treated for 4 days at 7.5 mg 2-14C pirimiphos-
methyl/kg bw/day and sacrificed at 24 h intervals did not show any
increase in blood levels with time. Tissue levels of total
radioactivity in liver, kidney and fat over the 4 day period were
generally below 2 mg pirimiphos-methyl equivalents/kg tissue (levels
of unchanged pirimiphos-methyl being less than 0.15 mg/kg tissue).
There was no evidence of tissue accumulation (Mills, 1976).
Adult male Wistar rats were intubated with 1 mg 14C
pirimiphos-methyl/kg bw/day. Four groups of 3 animals were dosed for
3, 7, 14 or 21 days and sacrificed 24 h after the final dose. A
further five groups of 3 rats were similarly dosed for 28 days and
sacrificed 1, 3, 7, 14, or 28 days post-dosing. Each of the 9 groups
had one rat, undosed, as a control. Following sacrifice, samples of
liver, kidney, muscle, fat, erythrocytes and plasma were taken for
analyses. Urine and faeces were collected from 2 rats over a 24 h
period following the seventh dose. Recovery of 14C from 14C
pirimiphos-methyl added to control tissues was 96.9±5.2%. In all
tissue samples at all time intervals, concentration of radioactivity
was very low, close to or below detection limits. Levels did not
increase with repeated dosing. Liver concentrations were fairly
constant (0.03 ppm) and in some kidney samples, similar levels were
detected. In other tissues, radioactivity concentration was
generally below the limits of detection (0.04-0.06 ppm). Three days
after cessation of dosing one animal had detectable levels of kidney
radioactivity concentrations. At 7 days and subsequent days, no
residues were found. Excretion was between 70 and 80% of a single
dose, following administration of 7 consecutive doses, providing
evidence of rapid metabolism and elimination rather than poor
absorption (Hawkins & Moore, 1979).
Dogs
Male beagle dogs (1/dose level) given 18.4 or 16.7 mg 2-14C-
ring labelled pirimiphos-methyl/kg bw by capsule excreted 64.4% or
82.5% of the administered dose in the urine, and 17.3% or 13.3% in
the faeces, respectively, in 48 h. Nine metabolites (unidentified)
were present in urine (Bratt & Dudley, 1970).
Goats
In the lactating goat, following a single dose of 0.12 mg 2-
14C-ring labelled pirimiphos-methyl/kg bw by capsule, 87% and 4%
were excreted in urine and faeces, respectively, over 8 days. 80.6%
and 2.7% were excreted in urine and faeces, respectively, during the
final 48 h. Milk residues during the first 24 h post-dosing
indicated a total residue of 0.035 ppm, of which only 0.003 ppm was
parent compound. After 24 h residues declined to 0.004 ppm or less
(Bowker et al., 1973).
A lactating female goat was orally dosed by gelatin capsule,
twice daily, with 40 mg 14C-pirimiphos-methyl (7.1 x 106ßg)
labelled in the 2 position of the pyrimidine ring, for 7 consecutive
days (equivalent to 45 ppm in the diet). Chemical and radiochemical
purity exceeded 99%. At sacrifice, 78.1 and 11.3% of total
administered radioactivity had been excreted in urine and faeces,
respectively. Residues in milk were approximately 0.2% of the total
administered dose, the highest residue being 0.208 ppm pirimiphos-
methyl equivalents at day 2 of dosing. Levels plateaued by day 4 at
about 0.15 ppm. Sixteen hours after sacrifice, radioactive residues
in liver, kidney, muscle and fat were 0.31, 0.5, 044, and 0.067 ppm,
respectively. The major components in fat were unchanged pirimiphos-
methyl (55.2% of total radioactive residue) and 0-(2-ethylamino-6-
methylpyrimidine-4-ol)0,0-dimethyl phosphorothioate (17.1%), and
those in the remaining tissues and in milk were 2-diethylamino-6-
methylpyrimidine-4-ol,2-ethylamino-6-methylpyrimidine-4-ol, and 2-
amino-6-methylpyrimidine-4-ol (Skidmore et al., 1985).
Cows
In the cow following a single oral dose of 0.5 mg 2-14C-
labelled pirimiphos-methyl/kg bw, excretion was similar to that in
goat, 85% and 14% of the administered dose being excreted in urine
and faeces, respectively, over 7 days. During the first 3 days post-
dosing, 0.35% of the labelled was excreted in the milk (residue
level, 0.04 ppm of which less than 2% was unchanged pirimiphos-
methyl and phosphorus-containing metabolites). Hydroxypyrimidine
hydrolyses products or their conjugates constituted the major
portion of the milk residues (Bullock et al., 1974).
Hens
Hens given a single oral dose of 2, 9 or 20 mg-14C-labelled
pirimiphos-methyl excreted over 70% of the administered dose within
24 h. Three major metabolites, namely parent pyrimidine (4.7% of
dose), 2-ethyl-amino-4-hydroxy-6-methyl pyrimidine (25% of dose) and
2-amino-4-hydroxy-6-methyl pyrimidine (31%) were identified. When
fed for 289 days with dietary concentrations of 4 ppm, levels of
parent compound in white and yolk of eggs never exceed 0.001 ppm
although total 14C levels increased to 0.032 to 0.038 ppm over 16
days, and then fell (to equilibrium) to 0.026 to 0.028 ppm, 86-90%
of which comprised water soluble metabolites. Hens fed dietary
concentrations of 32 ppm for 7 days yielded eggs containing 0.007
(whites on day 3) and 0.012 (yolks on day 6) ppm pirimiphos-methyl.
"Total" pirimiphos-methyl plus equivalents ranged from 0.08 to 0.15
ppm in whites plus yolk. In muscle, residues of 0.31 and 0.16 ppm
were present following exposure to 32 and 4 ppm dietary
concentrations, respectively (Green et al., 1973).
Two hens were dosed orally by gelatin capsule with a mean dose
of 1.52 mg 14C-pirimiphos-methyl (60.75 µ Ci)/hen for 14
consecutive days. Radiochemical purity was &gt 95%. Residues of 14C
in breast muscle were 0.59% and 0.39 ppm. No unchanged pirimiphos-
methyl was detected. The majority of the residue (74%) was
identified as free and conjugated 2-amino-6-methyl-pyrimidin-4-ol.
Radioactive residues in yolk appeared to plateau about days 7-10,
but residues in albumen were inconsistent. At least 7 different
compounds were identified as being present in the eggs, the major
ones being 2-ethylamino-6-methyl-pyrimidine-4 ol and 2-amino-6-
methyl-pyrimidine-ol (Hall et al., 1979).
14C-Pirimiphos-methyl, labelled at the 2 position of the
pyrimidine ring was administered to 3 hens by gelatin capsule, twice
daily for 14 days. The mean dose/day was 2.54 mg pirimiphos-methyl
(98.4% purity), equivalent to 50 ppm in the diet. An additional bird
was fed capsules treated similarly to those fed to experimental
birds, but containing no pirimiphos-methyl. Eggs were collected
daily and excreta at 24 h intervals. Sixteen hours after the final
dose, hens were killed, and tissues were taken for analysis. A mean
value of 97.5% of the administered dose was excreted over the 14 day
period. At sacrifice, liver, peritoneal fat, subcutaneous fat, leg
muscle and breast muscle contained 0.2, 0.093, 0.11, 0.67 and 1.3
ppm pirimiphos-methyl equivalents, respectively. In fat,
approximately 72% of the radioactive residue was unchanged
pirimiphos-methyl, which was only found elsewhere (9.5% of the total
radioactive residue) in egg yolk. In muscle tissue the major residue
was 2-amino-6-methylpyrimidin-4-ol and 2-ethylamino-6-
methylpyrimidine-4-ol. These compounds were also found in liver in
both free and conjugated forms (Skidmore & Tegala, 1985).
Biotransformation
The metabolism of pirimiphos-methyl in Wistar rats (5 males)
given 100 mg 14C-labelled pirimiphos methyl/kg bw and in one
beagle dog given 20 mg 14C-labelled pirimiphos-methyl/kg bw was
investigated by TLC separation of urinary metabolites. Twelve
metabolites were detected in rat, and 11 in dog urine. Five of the
metabolites were identified. In both species, 2-ethylamino-4-
hydroxy-6-methyl pyrimidine was the major urinary metabolite (30% of
dose). The next most predominant metabolite in dog was 4-0(2-
diethylamino-6-methylpyrimidinyl-ß-D-glucosiduronic acid (11% of
dose) and in the rat, an unidentified phosphorus-containing product
thought to be a dealkylated derivative of either pirimiphos-methyl
or its oxygen analogue (12% of dose). Other identified metabolites
were 2-amino-4-hydroxy-6-methyl pyrimidine (8% and 5% of dose in rat
and dog, respectively) (Green et al., 1973; Bratt & Jones, 1973).
These studies indicate that the P-O-C bond of pirimiphos-methyl
is extensively cleaved and that N-de-ethylation and/or conjugation
are further steps in the metabolism of the pyrimidine leaving group.
Although the oxygen analogue of pirimiphos-methyl was not detected
as a urinary metabolite, the fact that cholinesterase inhibition
occurs in vivo suggests that the oxygen analogue is also formed
and may be an intermediate step leading to the identified urinary
products (Annex 1, reference 23).
Effects on enzymes and other biochemical parameters
The only biochemical effects consistently noted in acute or
chronic toxicity tests was inhibition of cholinesterase. A group of
36 male rats were given single oral doses of 1450 mg pirimiphos-
methyl/kg bw. Symptoms were noted and they were sacrificed at
intervals up to 4 days after dosing for measurements of brain,
plasma and red cell cholinesterase activity. Few signs were noted at
6 h after dosing when cholinesterase inhibition was 0, 35 and 51%,
respectively, for brain, red cell and plasma. Clear signs of
poisoning only became apparent by 24 h when brain was inhibited by
46% and red cell and plasma by 70 and 80%, respectively. Recovery of
cholinesterase activity began to be apparent by 72 h. Plasma
cholinesterase activity had completely recovered by 96 h but red
cell and brain remained 47 and 30% inhibited, respectively, at this
time (Clark, 1970). From these studies it appears that inhibition of
brain cholinesterase by 40% or more results in obvious signs of
toxicity (Annex 1, reference 23).
A group of 25 male (probably Wistar CFT, but not specified)
were dosed with 1000 mg/kg bw. Twenty-five additional rats served as
controls. Five rats/group were sacrificed at 4, 8, 24, 48 or 72 h
post-dosing and plasma and brain cholinesterase and non-specific
carboxylesterase activities were measured. Plasma cholinesterase
inhibition was rapid (60% inhibition by 4 h) whereas brain
cholinesterase inhibition was slower (36% by 8 h). Both attained
maximum inhibition by 24 h (93% for plasma and 61% for brain).
Recovery was apparent at 48-72 h in both enzymes, but that for brain
was slower. Non-specific esterase (NSE) activity was inhibited,
attaining maximum inhibition (plasma 80%, and brain 47%) at 24 h.
Inhibition of NSE was less than for cholinesterase, and recovery in
plasma was more rapid. In brain, NSE and cholinesterase activity
recovery were comparable (Rajini & Krishnakumari, 1988a).
Pirimiphos-methyl, 90.5% purity, was fed at dietary
concentrations of 0, 1000 or 1500 ppm to groups of 30 male Wistar
rats for 28 days. Five rats/group were necropsied 7, 14, 21 or 28
days post-initiation of exposure, and 5 rats/treated group were
sacrificed at 35 days (i.e. after 7 days withdrawal from pirimiphos-
methyl exposure). The fate of the remaining 5 rats is not reported.
Brain and erythrocyte cholinesterase inhibition showed significant
dose-related depression at all time intervals during exposure.
Erythrocyte cholinesterase was consistent during exposure, but brain
cholinesterase did not achieve consistent levels until 14-21 days.
Post-exposure recovery occurred in all groups but in brain,
cholinesterase activity was still biologically significantly
depressed (26 and 28% at 1000 and 1500 ppm, respectively). Plasma
cholinesterase activity was variable, but was depressed 17-44% over
the various time intervals, the least depression being at the
highest dose. Recovery was complete 7 days after cessation of
dosing. Non-specific brain carboxylesterase activity was depressed
at 1500 ppm at all time intervals, but only after 14 days at 1000
ppm. Recovery was rapid and complete at both dose levels following
withdrawal. Plasma non-specific carboxylesterase activity was
markedly depressed at all time intervals, but was still
significantly depressed following 7 days withdrawal. Renal non-
specific carboxylesterase activity was slightly reduced only at 1500
ppm after 14 days treatment and recovered rapidly upon cessation of
dosing (Rajini et al., 1989).
Toxicological studies
Acute toxicity studies
Table 1. Acute toxicity of pirimiphos-methyl
Animal Sex Route LD50 (mg/kg) Reference
Mouse M oral 1180 (1030-1360) Clark, 1970
Rat F oral 2050 (1840-2260) Clark, 1970
Rat M oral 1861 (1266-2928) Rajini & Krishnakumari
(1988a)
Table 1 (cont'd)
Animal Sex Route LD50 (mg/kg) Reference
Rat F oral 1667 (1187-2284) Rajini & Krishnakumari
(1988a)
Guinea-pig F oral 1000-2000 Clark, 1970
Rabbit M oral 1150-2300 Clark, 1970
Cat F oral 575-1150 Clark, 1970
Dog M oral &gt 1500 Gage, 1972
Hen F oral 30-60 Clark, 1970
Quail F oral approx. 140 Gage, 1971a
Test material was 90-94% purity, with at least 14 days post-
dosing observation. Toxic signs were typical of those resulting from
cholinesterase inhibition.
Table 2. Acute toxicity of metabolites
Compound Animal Route LD50 (mg/kg) Reference
2-diethylamino-4-hydroxy-6- Rat oral 800-1600 Gage, 1971b
methylpyrimidine
2-ethylanino-4-hydroxy-6- Rat (F) oral 2093 (1841- Parkinson,
methylpyrimidine 2380) 1974
2-amino-4-hydroxy-6-methyl- Rat (F) oral &gt 4000 Parkinson,
pyrimidine 1974
Toxic signs reported for 2-ethylamino-4-hydroxy-6-methyl
pyrimidine comprise urinary incontinence and salivation. No toxic
signs were observed with 2-amino-4-hydroxy-6-methylpyrimidine.
Short-term toxicity studies
Rats
Repeated and daily dosing of 10 rats/sex/dose, 5 times weekly
for 2 weeks at 200 mg/kg bw/day produced mild signs of poisoning
(fibrillation and urinary incontinence) noted only after 7 doses and
not increasing in intensity. Body-weight gain was depressed;
haemoglobin levels were depressed about 9% and reticulocyte counts
were increased - marked aniscytosis and some anisochromia were noted
in erythrocytes as well as occasional nucleated erythrocytes and
some Howell-Jolly bodies; histopathological changes comprised
splenic haematopoiesis and in 1/8 animals, haemosiderosis. A second
identical study at 400 mg/kg bw/day resulted in signs of poisoning
in 2 days, increasing in severity and resulting in 9/10 deaths in
males, and 3/10 deaths in females. Haematology and histopathology
were similar to the effects noted at 200 mg/kg bw/day (Clarke,
1970).
Four groups of 25 Alderley Park SPF rats/sex/dose were fed
diets containing 0, 8, 80 or 360 ppm 93.1% purity pirimiphos-methyl
for 90 days. Twenty/sex/dose were sacrificed at termination of
dosing, and the remainder, 28 days later. Body-weight gain in
females was reduced 18 and 21%, compared to controls, at 80 and 360
ppm, but food intake in these groups was slightly increased. Plasma
cholinesterase was depressed in males (41-72%) and females (56-88%)
during weeks 2-12 at 80 and 360 ppm. Recovery to normal activity was
observed one week after withdrawal of pirimiphos-methyl. Erythrocyte
cholinesterase was depressed in males (39-52%) and females (43-71%)
at 360 ppm with incomplete recovery occurring by week one (40%
inhibition in males, and 30% inhibition in females). Brain
cholinesterase was depressed (mainly in females) at 80 and 360 ppm.
Recovery did not occur within the 4-week post-dosing period.
No effects were observed on haematological parameters
(haemoglobin concentration, PCV, MCHC, reticulocyte counts, total
and differential white cell counts, platelet counts, mean
corpuscular diameter and Kaolin-cephalin prothrombin time) or in the
incidence of gross or histopathological lesions relative to those
observed in controls. The NOAEL was 8 ppm, equivalent to 0.4 mg/kg
bw/day (Clapp and Conning, 1970).
Five groups of 12 young male rats (probably Wistar CFT strain,
but not specified) were fed dietary concentrations of 0, 10, 250,
500 or 1000 ppm pirimiphos-methyl (90.5% purity) for 28 days. There
were no effects on body-weight gain or food intake. Compound intake
was calculated to be 0, 4, 100, 200 or 400 mg/kg bw/day (data not
given). A slight increase in liver weight was reported at 1000 ppm.
No treatment-related pathological changes were observed in liver,
brain, lung, heart, adrenal, kidney, spleen or testes. Increased
serum transaminases (1000 ppm) and increased alkaline phosphatase
(500-1000 ppm) were noted. Hepatic transaminases (ß-glucoranidase
and alkaline phosphatase) were unaffected. Cholinesterase activity
(plasma and brain) were inhibited at 250 ppm and above. The NOAEL
appears to be 10 ppm (stated to be equal to 4 mg/kg bw) (Rajini &
Krishnakumari, 1988a).
Four groups of male Wistar CFT rats were fed dietary
concentrations of 0, 500, 1000 or 1500 ppm pirimiphos-methyl for 28
days. Four rats/group were sacrificed at 7, 14, 21 or 28 days.
Mortality, growth rate and food consumption were stated to be
unaffected at any dose level. Blood glucose levels in treated rats
were consistently below control levels. The authors stated that
"decrease in blood glucose level was evident at all dosages during
the second week and, at all intervals in 1500 ppm group". This
statement is questionable since, when converted to percentage of
control values, blood glucose level decrease was greatest in week 1
at 1000 (58%) and 1500 (61%) ppm. Dose-effect relationships are
difficult to determine and control values were high (125-138 mg/100
ml). Although standard deviations are given (3.06-9.20), in the
absence of individual data, interpretation is equivocal. Blood urea
levels were consistently elevated above control levels, but the
effect was generally greatest at the mid-dose. Dose/effect
relationships do not appear to be present, based on the data
available for evaluation. Similarly data on urinary excretion of
urea was sporadic and usually non-dose related. Protein excretion in
urine was generally increased but, except in week 4, dose/effect
relationships are questionable. Creatinine and creatine excretion is
equally difficult to interpret (Rajini & Krishnakumari, 1988b).
Four groups of 20 male Wistar CFT rats were administered 0
(coconut oil), 50, 100 or 200 mg 90.5% purity pirimiphos-methyl/kg
bw/day, five times weekly for 4 weeks. Four rats/group were
sacrificed on days 7, 14, 21 or 28. Signs of toxicity were seen only
in week 4 at 200 mg/kg bw/day. Body-weight and food intake were
comparable in all groups. Total RBC were depressed at all time
intervals and all doses except at 100 ppm and 3 weeks, which were
fractionally above control counts. Dose-effect relationships were
virtually non-existent despite statistically significant decreases
in mean values. A wide variation in control values (9.95 x 10-6/µl
- 8.05 x 10-6/µl) also render interpretation difficult, especially
with only 4 rats/group.
Similar problems existed in total white cell counts (15 275 -
28 500/µl in controls over the 4 week period) and differential
counts (12 800 - 23 400 for lymphocytes, 1800-4000 for neutrophils).
The only consistent effects seem to be the increased clotting time
and prothrombin time and decreased platelet counts seen at all dose
levels after week 2 (Rajini et al., 1987).
An additional short-term study in rats was reviewed. The
purpose of this study was to alleviate concerns that the rats used
in the long-term study were rather older (based on their body-weight)
than those normally used in such a study. Hence possible effects
(especially on cholinesterase activity and inhibition) may have
occurred in young rats. To ensure this was not the case, the
following study was conducted.
Groups of 12 rats/sex/dose were fed diets containing 0, 5, 8,
10 or 50 ppm 97% purity pirimiphos-methyl for 28 days. The rats were
approximately 6 weeks old at receipt and were placed on study over a
3-week period - females during the first week and males during the
third week. Plasma and erythrocyte cholinesterase activity were
measured on groups of 5 rats/sex/dietary concentration on days -14,
-7, 1, 3, 7, 14, 21 and 28 days, and brain cholinesterase was
measured on 5 rats/sex/dietary concentration on day 28. There were
no effects of pirimiphos-methyl on body-weight gain (animals weighed
weekly) or on clinical conditions and behaviour. Food consumption
(measured on groups of 3 rats/week) was reduced, males at 5 ppm
(statistically significant) and 8 ppm (not statistically
significant). At 5 ppm, this reduced food intake was associated with
a slight (non-statistically significant) decrease in body-weight
gain (ca. 5%). Food utilization was comparable in all groups. At
termination, gross pathology of rat(s)/sex/group did not reveal any
lesions attributable to pirimiphos-methyl. Plasma cholinesterase
depression consistently exceeded 20% in the 50 ppm group. Sporadic
inhibition was noted at 8 and 10 ppm, as was sporadic elevation.
Erythrocyte cholinesterase inhibition was unaffected by pirimiphos-
methyl even at 50 ppm. Brain choline-sterase inhibition exceeded 10%
in both sexes at 50 ppm but was not significantly affected at lower
dietary concentrations (Berry & Gore, 1975).
Dogs
Four groups of beagle dogs/sex were fed gelatin capsules at
pirimiphos-methyl doses of 0, 2, 10, or 25 mg/kg bw/day, for 13
weeks. Two dogs/sex/dose were sacrificed at 13 weeks, and the
remainder at 17 weeks. Clinical signs (dry skin, dull coat during
weeks 2 and 3, and increased incidence of vomiting during the first
5-6 weeks) were observed at 25 mg/kg bw/day. An increased frequency
of liquid stools was noted at 10 and 25 mg/kg bw/day. Body-weight
gain was reduced in both sexes at 25 mg/kg bw/day and in females at
10 mg/kg bw/day. At 2 mg/kg bw/day, female body-weight gain
reduction was of borderline significance. Reduced food intake (dose-
related) occurred in all female test groups and in males at 25 mg/kg
bw/day. At 2 and 10 mg/kg bw/day male food intake was inconsistent.
Heart rate was reduced (both sexes) at 25 mg/kg bw/day. Plasma
cholinesterase was depressed at all dose levels (more than 20%) but
recovery was rapid following cessation of dosing. Erythrocyte
cholinesterase activity was depressed in a dose-related pattern, the
onset being late (10-12 weeks) at 2 mg/kg bw/day. At 10 and 25 mg/kg
bw/day, inhibition increased with time. Terminal brain
cholinesterase activity at 13 and 17 weeks was comparable to
controls in all groups. Two dogs showed high ALAT and SAP levels
after 3 months dosing, and a third, a less marked ALAT increase at
25 mg/kg bw/day. Bile duct proliferation and portal cirrhosis
occurred in 1/2 males at 25 mg/kg bw/day, and bile duct
proliferation only in 1/2 males at 10 mg/kg bw/day at 13 weeks.
After withdrawal, minimal bile duct proliferation occurred in 1/2
dogs of each sex at 25 mg/kg bw/day. The NOAEL was 2 mg/kg bw/day,
based on reduced body weight gain in females at 10 mg/kg bw/day
(Noel et al., 1970).
Four groups of 4 beagle dogs/sex were dosed with 0, 0.5, 2 or
10 mg/kg bw/day via gelatin capsules for 2 years ( corn oil
solution). A 7-day break in dosing occurred at 140 days (due to
problems with liquefying the test material). Brain cholinesterase
activity was 81, 78 and 44% of control values at 0.5, 2 and 10 mg/kg
bw/day, respectively. Erythrocyte cholinesterase activity was
depressed with respect to pre-dosing levels at 2 mg/kg bw/day
(greater than 20% from week 12 onwards) and 10 mg/kg bw/day. Plasma
cholinesterase activity depression was 21-33% almost consistently
throughout the study at 10 mg/kg bw/day, 21-33% from week 4 and
frequently thereafter at 2 mg/kg bw/day, and infrequently (weeks 38,
77, and 102) by 20-25% at 0.5 mg/kg bw/day. One female dog died on
day 401 after showing few clinical signs of intoxication. At 10
mg/kg bw/day, reduced food intake and body-weight gain were noted.
Electrocardiogram records and ophthalmoscopy were normal in all
groups as were haematological parameters (erythrocyte counts, Hb,
PCV, HCHE, MCV, reticulocyte counts, total and differential white
cell counts, platelet counts and prothrombin index), and clinical
chemistry (urea, glucose, protein, SAP, K+ and Na+) except for a
slight increase in ALAT at 12, 26 and 38 weeks in some dogs at 10
mg/kg bw/day. Histopathological changes were comparable in all
groups although absolute and relative (to body-weight ) liver
weights were increased at 10 mg/kg bw/day (Rivett et al., 1973).
The 1974 JMPR (Annex 1, reference 23) did not appear to consider the
19% depression of brain cholinesterase activity at 0.5 mg/kg bw/day
to be toxicologically significant.
Subsequent data have been submitted (ICI, 1988) which comprise
historical control data on dog studies performed between 1970 and
1975. These data clearly indicate that the apparent brain
cholinesterase inhibition at 0.5 and 2 mg/kg bw/day are well within
historical control values, based on 8 studies from the same
laboratory, using the same measurement techniques. Concurrent
controls in the pirimiphos-methyl study were, in 2 males and 3
females, abnormally high. The Meeting concluded that the NOAEL was 2
mg/kg bw/day, based on reduced cholinesterase activity and reduced
body weight gain at 10 mg/kg bw/day.
The potential hepatic effect of pirimiphos-methyl on dog liver
was investigated using 8 male dogs dosed by capsule at 25 mg/kg
bw/day, and 2 male dogs dosed at 25 mg/kg bw/day for 8 weeks and
then increasing to 35 mg/kg bw/day (14 days), 45 mg/kg bw/day (7
days), and finally 50 mg/kg bw/day (17 days). Six control animals
received corn oil only. Four out of eight dogs at 25 mg/kg were
withdrawn from treatment for 2 days after week 3 because of toxic
signs. During week 4, 3/8 dogs were sacrificed (due to weight loss)
and during week 12, 1/2 dogs, after 7 days at 50 mg/kg bw/day was
taken off treatment for 5 days. Slight bile-duct proliferation was
seen in 2 dogs receiving 25 mg/kg bw/day for 13 weeks but not in any
other animals, even at 50 mg/kg bw/day. Investigation of plasma
alkaline phosphatase, plasma alanine aminotransferase, aspartate
aminotransferase, and plasma glutamate dehydrogenase showed
elevation of these enzymes in almost all test animals on occasions,
throughout the study. No correlation with histopathology was
apparent. Individual sensitivity was extremely variable (Garuti et al., 1976).
Long-term toxicity/carcinogenicity studies
Mice
Three groups of 52 CFLP mice/sex were fed dietary
concentrations of 0, 5 or 250 ppm 97.8% purity pirimiphos-methyl for
80 weeks. A fourth group was fed 300 ppm increasing weekly by 50 ppm
to a final concentration of 500 ppm. Satellite groups of 12 mice/sex
fed 0, 5 and 500 ppm were utilized for blood cholinesterase studies.
Mortality was not significantly affected. Body-weight gain was not
affected overall, although food consumption was slightly reduced in
females at 500 ppm over the 80 week period. Water consumption was
unaffected. Erythrocyte cholinesterase activity (measured in weeks
0, 12, 36 and 80) was depressed in males at 5 ppm in week 36 only,
and in both sexes at all time intervals at 500 ppm. Plasma
cholinesterase activity was significantly decreased (&gt 20%) at all
time intervals at 500 ppm, and at 5 ppm in males at weeks 12 and 36,
and in females at week 80. Histopathological changes (hepatocytic
vacuolation hepatic nodular hypoplasia in females only, renal foci
of lymphocytic infiltration, perivascular or peribronchial lymphoid
aggregations in the lung and abscesses and small haemorrhages in the
ovaries were not deemed to be of toxicological significance since
they occurred in all groups and no dose-relationship was apparent.
There was no evidence of tumour induction at dose levels up to 500
ppm pirimiphos-methyl. The NOAEL was 5 ppm, equal to 0.5 mg/kg
bw/day in male mice, and 0.6 mg/kg bw/day in female mice (Hunter et al., 1976).
Rats
Four groups of 49 Wistar SPF rats/sex were fed pirimiphos-
methyl at dietary concentrations of 0, 10, 50 or 300 ppm for 2
years. An additional 24 rats/sex/group were fed the same diets and
were sacrificed (8/sex/group) at 12, 26 and 52 weeks for brain
cholinesterase studies. At termination of dosing, 8 rats/sex/group
were fed control diet for 4-8 weeks prior to sacrifice, all other
rats were sacrificed at 2 years. Survival (33-56%) was unaffected by
pirimiphos-methyl at dose levels up to 300 ppm. Two outbreaks of
infection were reported - one at 16 weeks, when "several animals of
either sex developed swellings (lasting no more than 72 h) in the
area of the salivary glands". No changes in behaviour, appetite or
condition were observed. The condition, distributed between groups,
did not recur. The second outbreak, occurring in the latter weeks of
the test, was a respiratory disease, resulting in several deaths.
All surviving rats were treated with 18 mg oxytetracycline/kg bw/day
for 5 consecutive days during week 86.
Body-weight gain and food intake were comparable in all groups.
At 300 ppm, marked plasma cholinesterase inhibition (50-80%) and
some inhibition of erythrocyte and brain cholinesterase (20-40%)
were noted. The only other adverse effect at this dietary
concentration was slight anaemia in female rats. Only female rat
plasma cholinesterase was consistently inhibited (50-65%) at the 50
ppm dietary level. Brain cholinesterase was reduced (&gt 20%) in
males at weeks 26 and 104 at 50 ppm. Slight concurrent plasma
cholinesterase inhibition occurred in females at 10 ppm. Brain
erythrocyte cholinesterase was not affected at this dietary
concentration. Recovery of depressed cholinesterase activity was
usually complete following the 8-week withdrawal period. Organ
weights and weight ratios, and incidence of gross or
histopathological changes were comparable between groups. Tumour
incidences in treated groups were generally comparable to incidences
in control groups. The NOAEL was 10 ppm, equivalent to 0.5 mg/kg
bw/day (Gore et al., 1974a).
Reproduction studies
Rats
A multigeneration study (3 generations, 2 litters/generation
using b litters for parental animals) using groups of 24 female and
12 male Charles River CD rats fed diets containing 0, 20 or 200 ppm
nominal concentrations was performed. Since dietary analysis
indicated levels of only 3-8.7 ppm were, in fact, present in the
diet of the F0 generation (nominal level 20 ppm), the study at
this dose level was extended to produce a 4th generation (a and b
litters).
No adverse effects on parental mortality, body-weight, or gross
pathology were seen in any test group. Neither were toxic signs
observed. At 200 ppm, mating performance and pregnancy rate were
reduced in the production of F2-F3 litters and in F3 litters
at 20 ppm. The decreased pregnancy rate was dose-related. Litter
data (total litter losses, litter size, litter and mean pup weights,
pup mortality, and length of gestation) were comparable between
groups. Examination of the ultimate litters (F3b at 200 ppm and
F4b at 20 ppm) by organ weight analyses, skeletal staining of
10/pups/sex from each group, and histopathology of 10 pups/sex from
0 and 200 ppm dose levels did not reveal any compound-related
effects. A NOAEL was not demonstrated in this study (Palmer & James,
1972).
The testes of the F1b and F2b male rats were examined
histopathologically in an attempt to find an explanation for their
reduced mating performance. However, in the testes examined from
rats of the control and both treatment groups, the degree of
activity and maturity of the process of spermatogenesis were
comparable (Annex 1, reference 23).
Groups of 20 SPF CF strain rats/dose level were fed dietary
concentrations of 0, 5, 10 or 100 ppm continuously for 3 generations
(1 litter/generation). There were no consistent effects on parental
animals as assessed by toxic signs, mortality, food consumption,
body-weight, mating perfornance, pregnancy rate or duration of
gestation. Parental plasma and erythrocyte cholinesterase depression
(&gt 20%) was recorded in one or both sexes at 100 ppm after 7-9
weeks on diet (i.e. during the premating period). No effects were
observed in pups as determined by litter size, pup mortality, litter
or mean pup weights, gross pathology of F3a or incidence of
anomalies. The NOAEL was 100 ppm, equivalent to 5 mg/kg bw/day
(Palmer & Hill, 1976).
Special studies on potentiation
Combined administration of half LD50 dose of pirimiphos-
methyl with either gamma-BHC or dichlorvos to rats did not indicate
potentiation (Annex 1, reference 23). However, similar studies with
bioresmithrin indicated some possible potentiation but to an extent
which does not appear to be sufficient to be of practical
significance (Annex 1, reference 27).
Special study on mammalian cell transformation
An in vitro study utilizing Syrian hamster kidney cells
(fibroblast morphology, cell line BHK21/c13) exposed to doses
ranging from 0.23 to 2300 µg/ml, according to the method of Styles,
1977) failed to induce cell transformation (Trueman, 1983).
Special studies on teratogenicity
Rats
Three groups of 18-21 pregnant Alderby Park SPF rats were fed
diets of 0, 10 or 200 ppm from gestation days 1 through 20 (day of
insemination: day 0). Pups were removed by caesarean section on day
20. Maternal body-weight gain was slightly reduced on day 7 of
gestation, but was normal by day 20. No differences in food
consumption or gross pathology of the dams were observed.
Implants/dam, resorption incidence, sex ratio, and incidence of
malformations were within the normal range of distribution At 200
ppm, mean fetal weight was decreased, but this was probably a
consequence of a concomitant increase in litter size (Hodge and
Moore, 1972).
A second rat study used 24 Alpk:AP Wistar derived rats/dose
level. Rats were administered 0, 1.5, 15 or 150 mg pirimiphos-methyl
(purity 88.5% w/w, doses corrected for 90.9% purity) in corn oil/kg
bw/day by gavage on gestation days 7-16 inclusive (day 1 being the
day of confirmation of mating) indicated maternal toxicity (changes
in chemical signs and one death), decreased body-weight gain and
food intake were observed at 150 mg/kg bw/day. In the 1.5 mg/kg
bw/day pre-implantation losses were increased with concomitant
effects on numbers of live pups and mean gravid uterine and litter
weights. Values were, however, stated to be within normal ranges for
the strain. It is improbable that this observation is compound-
related because of the absence of similar findings at higher doses -
and the minimal exposure which may have occurred prior to
implantation. No compound- or dose-related effects on pup weight,
in utero survival or incidence of malformations/variants were
noted except for slight evidence of delayed ossification in pes
scores at 150 mg/kg bw/day. NOAELs for maternal toxicity (15 mg/kg
bw/day), embryotoxicity (15 mg/kg bw/day, based on reduced pes
scores) and teratogenicity (&lt150 mg/kg bw/day) were identified
(ICI, 1985).
Rabbits
Three groups of 16 or 17 artificially inseminated Dutch rabbits
were orally administered gelatin capsules containing corn-oil
solutions of pirimiphos-methyl to give 0, 1 or 16 mg/kg bw/day from
gestation days 1 to 28 inclusive. Three does died during the study;
two in the 1 mg/kg bw/day group, and one in the 16 mg/kg bw/day
group. Litter data, however, were only available in summary format,
on 10, 10 and 11 does at 0, 1 and 16 mg/kg bw/day respectively,
despite 16, 15 and 15 does surviving to term. Whether this
discrepancy is due to unsuccessful fertilization or pre-implantation
loss or total resorption of litters cannot be ascertained. Based on
the available data, mean number of implants/reported litter were
similar in all groups, and resorption rates were highest in the
control group. Litter size was increased at 16 mg/kg bw/day, and
mean litter weight was slightly depressed at this dose level. Sex
ratio (M/F) appears to increase with dose, but are stated (no data)
to be within normal ranges. One high-dose fetus, with placental
thrombosis and necrosis, showed multiple malformations. A dose-
related increase in the occurrence of 14 caudal vertebrae was noted.
Cholinesterase depression occurred in erythrocytes at 1 (23%) and 16
(32-60%) mg/kg bw/day and in plasma at 16 mg/kg bw/day (23%). The
NOAEL for embryofetal toxicity and teratogenicity was 16 mg/kg
bw/day. A NOAEL for maternal toxicity was not demonstrated since at
1 mg/kg bw/day, erythrocyte cholinesterase inhibition exceeded 20%
(Gore et al., 1974b).
Special studies on neurotoxicity
Hens
In a study designed to assess acute delayed neurotoxicity,
pirimiphos-methyl was administered by oral intubation as a solution
in arachid oil to groups of five Light-sussex adult hens at dose
levels of 20, 30, 40, 50 or 60 mg/kg bw. The LD50 was established
at approximately 79 mg/kg bw. Over an observation period of 21 days
post-dosing none of the animals showed signs of neurotoxicity. At
day 26 the surviving birds in the two highest dose groups (4/5 at 50
mg/kg bw and 2/5 at 60 mg/kg bw) were re-dosed after being protected
with intramuscular injection of atropine (10 mg/kg bw) and 2-PAM (50
mg/kg bw) and observed for a further 21 days. Group sizes were made
up to 5 with "new" protected birds which received a single dose of
pirimiphos-methyl. A further group of 5 birds were dosed with 500 mg
TOCP/kg bw as a positive control. Clinical observations were
confined to two birds in the 50 mg/kg bw dose group which showed
sporadic doubtful signs of incoordination. Surviving birds (2/5) in
60 mg/kg bw dose group showed no signs of neuropathological changes.
All birds in the positive control group showed signs of ataxia and
significant neuropathological changes. It is concluded that
pirimiphos-methyl does not cause delayed neurotoxicity following
acute doses up to 60 mg/kg bw (Annex I, ref. 27; Ross et al.,
1975).
In a study designed to examine the potential of pirimiphos-
methyl to cause delayed neurotoxicity after subchronic exposure,
groups of ten adult hens were dosed at 0 (untreated control), 0
(corn oil control), 0.5, 1.0, 2.5, 5.0 or 10 mg pirimiphos-methyl
(93.5% purity) kg bw/day by oral intubation for 90 days. A further
group of ten birds received 90 daily doses of 7.5 mg TOCP/kg bw as a
positive control. A further 2 groups of 10 hens received 90 doses of
either 5 or 10 mg pirimiphos-methyl/kg bw/day followed by a recovery
period of 90 days when the birds were not dosed. Dosing was
discontinued for birds in the two recovery groups when signs of
toxicity were severe and restarted when signs of recovery were
noted. The final recovery period was started when a bird had
received a total of 90 doses.
Analysis of test solutions indicated concentrations were within
8% of nominal values, and were stable up to 10 days. Mortalities
were 1/10, 3/10 and 4/10 in the main groups at 1.0, 2.5, 5.0 and 10
mg/kg bw/day; 1/10 and 4/10 in the 5 and 10 mg/kg bw/day ancillary
groups during treatment. Post-treatment deaths, during the recovery
period were 4/9 and 1/6 at 5 and 10 mg/kg bw/day, respectively.
Signs of toxicity, incidence and severity increasing with dose, were
observed at 1.0 mg/kg bw/day and above. There were no signs of
ataxia occurring as a result of delayed neurotoxicity except in the
positive control group (7/10 birds). Food intake was variable during
dosing, but overall indicated a dose-related decrease at 10 mg/kg
bw/day, and in the positive control group. Food intake increased in
the recovery groups after termination of dosing. Mean body-weight
values were decreased in one group at 5 mg/kg bw/day and both groups
at 10 mg/kg bw/day of dosing. Degenerative damages in nerve tissues
at histopathological examination were only observed in the positive
control group. The study did not indicate any evidence of delayed
neurotoxicity due to exposure of adult hens to toxic doses of
pirimiphos-methyl (Roberts et al., 1983).
A pilot study using 3 groups of 6 ISA hens protected with 50
mg/kg bw 2-PAM i.p. and 10 mg atropine sulphate/kg bw i.p. at dose
levels of 0, 100 or 117 mg pirimiphos-methyl/kg bw/p.o. with 14 day
post-dosing observation determined a dose of 100 mg pirimiphos-
methyl/kg bw to be a suitable dose for protected hens in a delayed
neuropathy assessment study.
Enzyme activity (NTE and AChE) were determined in hens given
corn oil (4 birds), TOCP (2 birds), protected hens given 100 mg
pirimiphos-methyl/kg bw (10 birds), p.o. with sacrifice at 24 h (2
control, 1 TOCP and 3 treated birds) and 48 h (similar numbers). NTE
was unaffected in control and pirimiphos-methyl dosed birds, but
markedly inhibited in those given TOCP at both 24 and 48 h.
Conversely, TOCP had minimal effect on brain or spinal cord
acetylcholinesterase, which was markedly inhibited by pirimiphos-
methyl.
In the main study, 10 hens were given corn oil, 10 TOCP, and 40
received 100 mg pirimiphos-methyl in corn oil/kg bw p.o., the dose
being repeated after 21 days in control and pirimiphos-methyl
treated birds. TOCP birds were sacrificed at 21 days, and the others
at 42 days. Delayed ataxia was only seen in TOCP treated birds.
Control and pirimiphos-methyl treated birds showed similar spinal
cord and peripheral nerve histopathology, whereas TOCP showed
disruption, fragmentation and distortion of some spinal cord axon
with some damage to the myelin shealt or peripheral nerves as well
as axonal degeneration (Lock & Johnson, 1990; Hawkin et al.,
1989).
Special studies on genotoxicity
The results of genotoxicity studies on pirimiphos-methyl are
summarized in Table 3.
Table 3. Results of genotoxicity assays on pirimiphos-methyl
Test Dose Result Reference
Male mouse dominated 15, 80 or 150 mg/kg bw/day Pregnancy frequency McGregor, 1975
lethal for 5 days prior to pairing reduced at 150 mg/kg bw
Salmonella typhimurium ditto + Hanna & Dyer, 1975
(No metabolic activation)
Escherichia coli ditto - Hanna & Dyer, 1975
(No metabolic activation)
Micronucleus ditto - Seiber (1975)
Micronucleus (mouse) 100, 200 and 400 mg/kg bw - Rajini et al. (1986)
Sperm-morphology 100, 200 and 400 mg/kg bw - Rajini et al. (1986)
(mouse)
Bone marrow cytogenetic Single dose - Done & McGregor (1980)
study (rat) 32, 102, 320 mg/kg bw
Bone marrow cytogenetic 5 consecutive daily - ve at 32 and 102 mg/kg Done & McGregor (1980)
study (rat) 32, 102, 320 mg/kg bw bw. At 320 mg/kg
bw/day, chromatid gap
incidence was increased
and one chromosome gap
was recorded
Salmonella typhimurium 1.6-5000 µg/plate - Callander (1984)
TA1535, TA1537, TA1538,
TA98 and TA100 (with
and without metabolic
activation)
Table 3 (Cont'd)
Test Dose Result Reference
Mouse lymphoma cell 12.5-200 µg/ml - Cross (1986)
assay (L5178Y cells) (with
and without metabolic
activation
Human lymphocyte
cytogenetics (with and
without metabolic - Wildgoose et al. (1986)
activation) -
DONOR 1 12, 58, 116 µg/ml**
DONOR 2 12, 29, 58 µg/ml**
Hamster sister chromatid
exchange in lung
fibroblasts +- *** Howard et al. (1986)
with metabolic activation 0.14, 0.29, 1.4, 2.9, 14, 29 +-****
without metabolic 0.14, 0.29, 1.4, 2.9, 14, 29,
activation 145 µg/ml
* The doses in this study tended to be low and metaphase chromosome spread was variable, resulting in
reduced numbers of cells available for scoring.
** Maximum tolerated doses for the different donors.
*** Positive results (dose-related) were noted at 14, 29 and 145 µg/ml. At 145 µg/ml, the assessment was
based on a single cellulite at a dose which exhibited marked toxicity.
**** Positive results were noted at all dose levels except 2.9 µg/ml. Dose/effect relationships were not
clearly established.
Observations in humans
A dose of 0.25 mg 97.8% purity pirimiphos-methyl/kg bw/day was
taken orally for 28 days by 5 healthy males (59.5-73 kg bw, 25-45
years old). Blood samples were taken on days -14, -7, 1, 3, 7, 14,
21 and 28. One subject showed plasma cholinesterase inhibition
(21.5%) on day 28. Otherwise variations, both above and below pre-
dosing values, were within 12%. Four of five subjects had red-cell
cholinesterase activity values slightly below the pre-exposure
values during the last 2 weeks of the study. However, the group
means for each time interval did not differ significantly and the
variations noted were within the range of variations found by others
for normal untreated subjects (Chart et al, 1974).
Three male (62-73 kg, 22-27 years old) and 4 females (44-60 kg,
21-49 years old) were given 0.25 mg 97.8% purity pirimiphos-
methyl/kg bw/day by capsule for 56 days. Blood samples were taken
twice prior to initiation of dosing, and on days 7, 14, 21, 28, 35,
42, 49 and 56 of the study and also in the recovery period 7, 14, 21
and 29 post-treatment. Controls comprised 2 females (44 and 46 kg,
aged 29 and 30). No compound-related effects were observed on liver
function (alanine aminotransferase, aspartate aminotransferase,
alkaline phosphatase and glutanyl transpeptidase in plasma),
haematology (Hb, PCV, MCHC, total and differential WBC, platelets,
ESR) or erythrocyte cholinesterase. Plasma cholinesterase was
depressed about 20% in 2/4 females on days 14, 21 and 28 and in one
female on days 28 and 35. The effect did not increase with time. All
values were normal during the withdrawal period (Howard & Gore,
1976).
Two groups of spray workers (5/group) together with 1 mixer-
loader per group applied actellic 50 EC to grain using a compression
sprayer, hand held, for 3 consecutive days (4-5 h/day). The
formulation, containing 50% pirimiphos-methyl, was sprayed at a
concentration of 5 g/l with an application rate of 0.158 a.i./m2.
A second study, using similar groups applied actellic 40 EC
containing 40% pirimiphos-methyl sprayed at a concentration of 46
g/l and application rate of 2.0 a.i./m2 in unoccupied dwellings
for mosquito control. In the two studies, spray workers in one group
were in normal clothing, in the second group, protective clothing
(cotton overalls, caps, canvas shoes and safety glasses, and in the
case of the dwelling spray workers, cotton masks). Mixer-loaders
wore rubber boots, gloves, caps, cotton jackets, cotton masks and
safety glasses. Cholinesterase (blood and plasma) was measured on 3
consecutive days prior to spraying, at lunchtime and the evening on
the days of spraying, and on days 1, 2, 3 and 10 post-spraying.
Clinical examinations were performed by registered practitioners and
comprised case histories and a general medical examination with
emphasis on gastrointestinal, neuromuscular, cardiorespiratory,
visual and psychological effects and peripheral and central nervous
systems prior to, during, and subsequent to spraying. Cholinesterase
depression occurred on a group basis in all groups over the spray
period, with full recovery within 24 h post-dosing. The greatest
depression occurred in workers applying the 40 EC formulation. Ten
of the 24 exposed individuals showed &gt 15% depression of
erythrocyte cholinesterase, the maximum depression being 23%. Levels
of plasma cholinesterase inhibition were inconsistent, and less than
those seen for erythrocyte cholinesterase inhibition. There were no
clinical manifestations of toxicity (Chester & Hart, 1986).
COMMENTS
Following oral administration of pirimiphos-methyl to male
rats, 80.7% and 7.3% of the administered dose were excreted via
urine and faeces, respectively, within 24 h. In the dog, 48 h after
dosing with either 18.4 or 16.7 mg/kg bw, urinary excretion was
64.4% or 82.5% and faecal excretion 17.3% or 13.3%.
Metabolic data indicated that the P-O-C bond of pirimiphos-
methyl was readily cleaved and that N-de-ethylation and/or
conjugation were further steps in the metabolism of the pyrimidine
leaving group. Although the oxygen analogue of pirimiphos-methyl was
not detected as a urinary metabolite, the fact that cholinesterase
inhibition occurred in vivo suggests that the oxygen analogue was
also formed and may be an intermediate step leading to the
identified urinary products.
In rats and dogs 2-ethylamino-4-hydroxy-6-methylpyrimidine was
the major metabolite (30% of the administered dose).
The oral toxicity of pirimiphos-methyl is low. WHO has
classified the compound as slightly hazardous (WHO, 1992).
The only biochemical effect consistently observed with
pirimiphos-methyl in acute short-term or long-term studies was
cholinesterase inhibition.
In a series of short-term rat studies at dose levels of 0, 8,
80 or 360 ppm for three months, 0, 10, 250, 500 or 1000 ppm for 28
days, 200 mg/kg bw five times weekly for 14 days, and 0, 5, 8, 10 or
50 ppm for 28 days (young rats) the overall NOAEL was 10 ppm
(equivalent to 0.5 mg/kg bw/day) with effects on erythrocyte
cholinesterase and brain acetylcholinesterase at 80 ppm. At high
dose levels (200 mg/kg bw, five times weekly for two weeks)
erythrocyte morphology was affected. The NOAEL in young rats was
also 10 ppm, with brain (but not erythrocyte) acetylcholinesterase
depressed at 50 ppm after 28 days.
In two dog studies (13 weeks at dose levels of 0, 2, 10 or 25
mg/kg bw/day via capsule and 0, 0.5, 2 or 10 mg/kg bw/day for two
years by capsules) the NOAEL was 2 mg/kg bw/day, based on brain
acetylcholinesterase inhibition.
In an 80-week study in mice at dietary concentrations of 0, 5,
250 or 500 ppm, a NOAEL based on blood cholinesterase depression was
5 ppm (equal to 0.5 mg/kg bw/day) (blood cholinesterase was not
measured at 250 ppm, only at 5 and 500 ppm). Pirimiphos-methyl was
not carcinogenic in mice.
In a two-year study in rats at dietary concentrations of 0, 10,
50 or 300 ppm, tumour incidence was comparable to controls. The
NOAEL was 10 ppm (equivalent to 0.5 mg/kg bw/day) with brain
acetylcholinesterase inhibition occurring at higher levels.
Pirimiphos-methyl was not carcinogenic in rats.
In a four-generation reproduction study in rats at nominal
dietary concentrations of 0, 20 or 200 ppm, dose-related reduction
of pregnancy rates and reduced mating performance at 200 ppm were
noted. Dietary analyses indicated that the 20 ppm diet only
contained 9 ppm pirimiphos-methyl. No NOAEL was demonstrated in this
study.
A repeat study at dietary concentrations of 0, 5, 10 or 100 ppm
for three-generations (1 litter/generation) did not show any adverse
effects on reproductive parameters at any dose level. The NOAEL was
100 ppm (equivalent to 5 mg/kg bw/day) for reproductive effects.
In two rat teratology studies, one at dietary concentrations of
0, 10, or 200 ppm and the second at dose levels of 0, 1.5, 15, or
150 mg/kg bw/day, dosing extending over or beyond the period of
embryogenesis did not demonstrate any evidence of teratogenicity.
Fetotoxicity was observed at 200 ppm (equivalent to 10 mg/kg bw/day)
and 150 mg/kg bw/day. NOAELs for maternal toxicity (15 mg/kg
bw/day), embryotoxicity (15 mg/kg bw/day) and teratogenicity
(&lt 150 mg/kg bw/day) were identified.
A rabbit teratogenicity study at doses of 0, 1 or 16 mg/kg
bw/day administered from days 1-28 of gestation did not show any
evidence of teratogenic effects. The NOAEL for fetotoxicity and
teratogenicity was 16 mg/kg bw/day.
Four studies in hens indicated that pirimiphos-methyl does not
cause delayed neurotoxicity.
After considering the available in vitro and in vivo
genotoxicity data, the Meeting concluded that pirimiphos-methyl was
not genotoxic.
In two experimental studies with human volunteers of 28 and 56
days, the highest dose tested in both studies (0.25 mg/kg bw/day)
failed to induced erythrocyte cholinesterase inhibition in either
study.
In determining the ADI the first multigeneration study in rats
was discarded because the dietary concentrations were uncertain, and
the adverse effects noted (decreased pregnancy rate and mating
performance) were atypical of those normally seen in reproduction
studies with organophosphorous esters (decreased pup weight gain and
pup mortality during early lactation). A clear NOAEL of 100 ppm
(equivalent to 5 mg/kg bw/day, the highest dose tested) was
demonstrated in the repeat study.
Studies with mice, rats, and dogs, showed NOAELs of 0.5 mg/kg
bw/day or above. In human studies, no cholinesterase inhibition was
seen at 0.25 mg/kg bw/day (the highest dose tested). On this basis,
the Meeting revised the ADI to 0.03 mg/kg bw/day by applying a 10-
fold safety factor to the NOAEL in the human studies.
TOXICOLOGICAL EVALUATION
Level causing no toxicological effect
Mouse: 5 ppm, equal to 0.5 mg/kg bw/day (80-week study)
Rat: 10 ppm, equivalent to 0.5 mg/kg bw/day (two-year
study)
100 ppm, equivalent to 5 mg/kg bw/day (three-
generation reproduction study)
Dog: 2 mg/kg bw/day (two-year study)
Humans: 0.25 mg/kg bw/day
Estimate of acceptable daily intake for humans
0-0.03 mg/kg bw
Studies which will provide information valuable in the continued
evaluation of the compound
Further observations in humans.
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